基于部分rpoB基因序列分析的变形杆菌的系统发育树

Hussein Ali Mutlag, Intesar N. Khelkal
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摘要

由于提出的rpoB基因作为微生物群落研究的替代生物标志物的重要性,本研究将重点放在本地变形杆菌临床分离株的系统发育关系上。收集50株分离菌,通过培养、镜检和生化试验进行表型鉴定。使用VITEK 2紧凑型系统进行验证。选取10株分离菌株,提取DNA,用rpoB基因特异性引物扩增,凝胶电泳鉴定基因型。产物以1090 bp的分子大小带进行检测,使用ABI3730XL自动dna测序仪进行Sanger测序,并与GenBank中的标准序列进行分析和比较。分离株已在美国国家生物技术信息中心(NCBI)登记,编号为HIMUS1、HIMUS2、HIMUS3、HIMUS4、HIUS5、HIMUS6、HIMUS7、HIMUS8、himus9和HIMUS10。利用rpoB基因部分序列(895 bp)构建了系统发育树。采用geneous软件,用最大复合似然法计算了1000次自举重复的进化距离。与NCBI标准株序列的相似性在98.76% ~ 100%之间。关键词:rpoB基因,变形杆菌,测序,系统发育分析
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Phylogenetic tree of Proteus spp. Based on partial rpoB gene sequence analysis
Due to the importance of the proposed rpoB gene as an alternative biomarker for microbial community studies, this study has focused on phylogenetic relationships among local Proteus clinical isolates. Fifty bacterial isolates were collected and identified phenotypically according to the culture, microscopic examination and biochemical tests. VITEK 2 compact system was used to confirm identification. Genotypic identification was performed after DNA extraction for 10 selected isolates and amplification with rpoB gene-specific primer and gel electrophoresis. The products were detected with a ( 1090 bp ) molecular size band, which was sent for Sanger sequencing using an ABI3730XL automated DNA sequencer, and data were analyzed and compared with standard sequences in GenBank.The isolates have been registered in the National Center for Biotechnology Information (NCBI) with accession numbers and named (HIMUS1, HIMUS2, HIMUS3, HIMUS4, HIUS5, HIMUS6, HIMUS7, HIMUS8, HIMUS9 and HIMUS10 ). The phylogenetic tree was constructed using partial (895 bp) rpoB gene sequences for those ten strains. Evolutionary distances were calculated using the method of Maximum Composite Likelihood with 1000 bootstrap replicates using GENEIOUS software. The sequences presented a similarity percentage ranging between (98.76% and 100%) when compared with the sequences of standard strains in NCBI. Keywords: rpoB gene, Proteus spp. Sequencing, Phylogenetic analysis
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